The invention relates to tissue and organ transplantation.
The field of organ transplantation has enjoyed substantial progress during the last two decades, resulting in marked improvements in short-term graft survival. Organ transplant recipients, however, still face substantial risks of long-term morbidity and mortality. Though modern immunosuppressive regimens have led to a dramatic reduction of the incidence of acute rejection episodes, they have yet to achieve a similar effect for chronic rejection, which is still the leading cause of graft loss during long-term follow-up. In addition, the requirement for life-long immunosuppressive drug therapy carries a significant risk of severe side effects, including tumors, infections and metabolic disorders. The reliable induction of donor-specific tolerance would solve both problems by obviating the need for chronic non-specific immunosuppression and by abrogating detrimental immunological reactions against the allograft.
The invention provides methods of inducing tolerance to foreign antigens. The methods feature preparative regimens which may minimize or eliminate the need for thymic irradiation, T cell inhibiting antibodies, and in some cases hematopoietic space-creating irradiation, e.g. preparative whole body irradiation.
Accordingly, the invention features a method of promoting acceptance, by a recipient mammal, of a graft from a donor mammal of a second species. The method includes:
administering to the recipient, an inhibitor, e.g., a blocker, of a costimulatory pathway, (e.g., one or both of, an inhibitor, e.g., a blocker, of the CD40 ligand-CD40 interaction and an inhibitor, e.g., a blocker, of the CD28-B7 interaction);
introducing, e.g., by intravenous injection, into the recipient mammal, hematopoietic stem cells, e.g., a bone marrow preparation; and
preferably, implanting the graft in the recipient. The hematopoietic cells are believed to prepare the recipient for the graft that follows, by inducing tolerance at both the B-cell and T-cell levels.
In preferred embodiments the CD40 ligand-CD40 interaction is inhibited by administering an antibody or soluble ligand or receptor for the CD40 ligand or CD40, e.g., by administering an anti-CD40L antibody, e.g., 5c8 or an antibody with similar efficacy or an antibody whose epitope overlaps that of 5c8, (see U.S. Pat. No. 5,474,711, hereby incorporated by reference). Preferably the inhibitor binds the CD40 ligand.
In preferred embodiments the CD28-B7 interaction is inhibited by administering a soluble ligand or receptor or antibody for the CD28 or B7, e.g., a soluble CTLA4, e.g., a CTLA4 fusion protein, e.g., a CTLA4 immunoglobulin fusion, e.g, CTLA4/Ig. Preferably, the inhibitor binds B7. In preferred embodiments anti-B7-1 and/or anti-B7-2 antibodies are administered.
In preferred embodiments CTLA4-Ig and an anti-CD40L antibody are administered.
In preferred embodiments, a blocker of the CD40/CD40L interaction, e.g., an anti-CD40L antibody is administered prior to administration of a blocker of the CD28/B7 interaction, e.g., CTLA4/Ig. The CD40/CD40L blocker can be administered on the day donor tissue is introduced and the CD28/B7 blocker administered 2, 3,4 5 or more days later.
The recipient mammal can be, by way of example, a human. The donor mammal can be, by way of example, a swine, e.g., a miniature swine. The graft is preferably from a discordant species. The graft preferably expresses a major histocompatibility complex (MHC) antigen, preferably a class II antigen. In particularly preferred embodiments the recipient is a primate, e.g., a human, and the donor is a swine, e.g., a miniature swine.
In preferred embodiments the method can be practiced without the administration of hematopoietic space-creating irradiation, e.g., whole body irradiation.
In certain embodiments the method is practiced without T cell depletion or inactivation, e.g., without the administration of thymic irradiation, or anti-T cell antibodies.
In certain embodiments the method is practiced with T cell depletion or inactivation, e.g., by the administration of thymic irradiation, or anti-T cell antibodies.
In certain embodiments the method is practiced with partial T cell depletion or inactivation, e.g., by the administration of thymic irradiation, or anti-T cell antibodies, in such amount to result in partial depletion of recipient T cells.
In preferred embodiments the method includes administering a sufficiently large number of donor hematopoietic cells to the recipient such that, donor stem cells engraft, give rise to mixed chimerism without space-creating treatment. Thus, in preferred embodiments inhibitors of both pathways and a quantity of hematopoietic stem cells sufficient to give rise to mixed chimerism, without the need for hematopoietic space-creating irradiation, is administered to the recipient. In preferred embodiments the number of donor hematopoietic stem cells is at least 200%, is at least equal to, or is at least 75, 50, or 25% as great as, the number of bone marrow hematopoietic stem cells found in an adult of the recipient species. In the case where an inbred population of the donor species exists, e.g., where the donor species is miniature swine, the number of available donor cells is not limited to the number of cells which can be obtained from a single animal. Thus, in such cases, the donor cells administered to the recipient can come from more than one, e.g., from two, three, four, or more animals. As is discussed below the donor stem cells can be provided in two or more separate administrations.
In preferred embodiments, mixed chimerism is induced in the recipient and the state of mixed chimerism is formed in the absence of the induction of hematopoietic space, e.g., in the absence of hematopoietic space created by space creating irradiation, e.g., whole body irradiation.
The number of donor cells administered to the recipient can be increased by either increasing the number of stem cells provided in a particular administration or by providing repeated administration of donor stem cells.
Repeated stem cell administration can promote engraftment, mixed chimerism, and preferably long-term deletional tolerance in graft recipients. Thus, the invention also includes methods in which multiple hematopoietic stem cell administrations are provided to a recipient. Multiple administration can substantially reduce or eliminate the need for hematopoietic space-creating irradiation. Administration can be given prior to, at the time of, or after graft implantation. In preferred embodiments multiple administrations of stem cells are provided prior to the implantation of a graft. Two, three, four, five, or more administrations can be provided. The period between administrations of hematopoietic stem cells can be varied. In preferred embodiments a subsequent administration of hematopoietic stem cell is provided: at least two days, one week, one month, or six months after the previous administration of stem cells; when the recipient begins to show signs of host lymphocyte response to donor antigen; when the level of chimerism decreases; when the level of chimerism falls below a predetermined value; when the level of chimerism reaches or falls below a level where staining with a monoclonal antibody specific for a donor PBMC antigen is equal to or falls below staining with an isotype control which does not bind to PBMC""s, e.g. when the donor specific monoclonal stains less than 1-2% of the cells; or generally, as is needed to maintain a level of mixed chimerism sufficient to maintain tolerance to donor antigen.
One or more post graft-implantation-administrations of donor stem cells can also be provided to minimize or eliminate the need for irradiation. Post graft administration of hematopoietic stem cells can be provided: at least two days, one week, one month, or six months after the previous administration of stem cells; at least two days, one week, one month, six months, or at any time in the life span of the recipient after the implantation of the graft; when the recipient begins to show signs of rejection, e.g., as evidenced by a decline in function of the grafted organ, by a change in the host donor specific antibody response, or by a change in the host lymphocyte response to donor antigen; when the level of chimerism decreases; when the level of chimerism falls below a predetermined value; when the level of chimerism reaches or falls below a level where staining with a monoclonal antibody specific for a donor PBMC antigen is equal to or falls below staining with an isotype control which does not bind to PBMC""s, e.g. when the donor specific monoclonal stains less than 1-2% of the cells; or generally, as is needed to maintain tolerance or otherwise prolong the acceptance of a graft.
When multiple stem cell administrations are given one or more of the administrations can include a number of donor hematopoietic cells which is at least 200%, is equal to, or is at least 75, 50, or 25% as great as, the number of bone marrow cells found in an adult of the recipient species; include a number of donor hematopoietic stem cells which is at least 200%, is equal to, or is at least 75, 50, or 25% as great as, the number of bone marrow hematopoietic stem cells found in an adult of the recipient species.
Although methods in which blockers of both pathways are administered may usually minimize or eliminate the need for other preparative steps, some embodiments include inactivating natural killer cells, preferably graft reactive or xenoreactive, e.g., swine reactive, NK cells, of the recipient mammal. This can be accomplished, e.g., by introducing into the recipient mammal an antibody capable of binding to natural killer cells of the recipient mammal, e.g., an anti-CD2 antibody, e.g., MEDI-507. The administration of antibodies, or other treatment to inactivate natural killer cells, can be given prior to introducing the hematopoietic stem cells into the recipient mammal or prior to implanting the graft in the recipient. This antibody can be the same or different from an antibody used to inactivate T cells.
Although methods in which blockers of both pathways are administered may usually minimize or eliminate the need for other preparative steps, some embodiments include inactivating T cells, preferably graft reactive or xenoreactive, e.g., swine reactive, T cells of the recipient mammal. This can be accomplished, e.g., by introducing into the recipient mammal an antibody capable of binding to T cells of the recipient mammal. The administration of antibodies, or other treatment to inactivate T cells, can be given prior to introducing the hematopoietic stem cells into the recipient mammal or prior to implanting the graft in the recipient. This antibody can be the same or different from an antibody used to inactivate natural killer cells.
One source of anti-NK antibody is anti-human thymocyte polyclonal anti-serum. Preferably, a second anti-mature T cell antibody can be administered as well, which lyses T cells as well as NK cells. Lysing T cells is advantageous for both bone marrow and graft survival. Anti-T cell antibodies are present, along with anti-NK antibodies, in anti-thymocyte anti-serum. Repeated doses of antibodies, e.g., anti-NK or anti-T cell antibodies, may be preferable. Monoclonal preparations can be used in the methods of the invention. An anti-CD2 antibody, e.g., MEDI-507, can be used as an anti-NK antibody.
Other preferred embodiments include: the step of introducing into the recipient mammal, donor species-specific stromal tissue, preferably hematopoietic stromal tissue, e.g., fetal liver or thymus. In preferred embodiments: the stromal tissue is introduced simultaneously with, or prior to, the hematopoietic stem cells; the hematopoietic stem cells are introduced simultaneously with, or prior to, the antibody.
Although methods in which blockers of both pathways are administered may usually minimize or eliminate the need for other preparative steps, some embodiments include the inactivation of thymocytes or T cells which can be performed prior to hematopoietic stem cell or graft transplantation. In preferred embodiments the method includes diminishing or inhibiting thymocyte or T cell activity, preferably the activity of thymic or lymph node T cells by administering to the recipient a short course of an immunosuppressive agent, e.g., a chemical or drug, e.g., cyclosporine, sufficient to inactivate thymocytes or T cells, preferably thymic or lymph node T cells. The duration of the short course of immunosuppressive agent is: approximately equal to or less than 30, 40, 50, 60, 120, or 365 days; approximately equal to or less than 8-12 days, preferably about 10 days; approximately equal to or less than two, three, four, five, or ten times the 8-12 or 10 day period. The short course can begin: before or at about the time the treatment to induce tolerance is begun, e.g., at about the time stem cells are introduced into the recipient; on the day the treatment to induce tolerance is begun, e.g., on the day stem cells are introduced into the recipient; within 1, 2, 4, 6, 8, 10, or 30 days before or after the treatment to induce tolerance is begun, e.g., within 1, 2, 4, 6, 8, 10, or 30 days before or after stem cells are introduced into the recipient. The short course of an immunosuppressive can be administered in conjunction with an anti-T cell antibody The short course of an immunosuppressive should be sufficient in concentration and duration to inactivate T cells, e.g., thymic or lymph node T cells, which would not be inactivated by antibody-based inactivation of T cells, e.g., inactivation by intravenous administrations of ATG antibody, or similar, preparations.
Although methods in which blockers of both pathways are administered may usually minimize or eliminate the need for other preparative steps, some embodiments include (optionally): the step of, prior to hematopoietic stem cell transplantation, creating hematopoietic space, e.g., by irradiating the recipient mammal with low dose, e.g., less than 400, preferably less than 300, more preferably less than 200 or 100 cGy, whole body irradiation to deplete or partially deplete the bone marrow of the recipient. As is discussed herein this treatment can be reduced or entirely eliminated. Other methods of creating hematopoietic space, e.g., administering hematopoietic space creating antibodies or drugs, e.g., cyclophosphamide or busulfan, to the recipient, can be used. E.g., hematopoietic space can be formed by administering an inhibitor of cell proliferation, e.g., DSG, or an anti-metabolite, e.g. brequinar, or an anti-T cell antibody, e.g., one or both of an anti-CD4 or anti-CD8 antibody.
Other preferred embodiments include: the step of, preferably prior to hematopoietic stem cell transplantation, depleting natural antibodies from the blood of the recipient mammal. Depletion can be achieved, by way of example, by contacting the recipients blood with an epitope which absorbs preformed anti-donor antibody. The epitope can be coupled to an insoluble substrate and provided, e.g., as an affinity column. E.g., an xcex11-3 galactose linkage epitope-affinity matrix, e.g., matrix bound linear B type VI carbohydrate, can be used to deplete natural antibodies. Depletion can also be achieved by hemoperfusing an organ, e.g., a liver or a kidney, obtained from a mammal of the donor species. (In organ hemoperfusion antibodies in the blood bind to antigens on the cell surfaces of the organ and are thus removed from the blood.)
Other preferred embodiments include those in which: the same mammal of the second species is the donor of one or both the graft and the hematopoietic cells; and the antibody is an anti-human thymocyte polyclonal anti-serum, obtained, e.g., from a horse or pig.
In preferred embodiments, the method includes the step of introducing into the recipient a graft obtained from the donor which is obtained from a different organ than the hematopoietic stem cells, e.g., a heart, pancreas, liver, or kidney.
In a particularly preferred embodiment the method includes:
administering to a human recipient one or both, of an inhibitor, e.g., a blocker, of the CD40 ligand-CD40 interaction and an inhibitor, e.g., a blocker, of the CD28-B7 interaction;
introducing, e.g., by intravenous injection, into the recipient mammal, hematopoietic stem cells, e.g., a bone marrow preparation; and
implanting the graft in the recipient. The method can be practiced without T cell depletion or inactivation, with T cell depletion or inactivation, or with partial T cell depletion or inactivation. T cell inactivation can be effected by the administration of thymic irradiation, or anti T cell antibodies.
In preferred embodiments donor tissue, e.g., hematopoietic cells, are depleted, e.g., partially or wholly, of donor T cells.
In another aspect, the invention features a method of promoting acceptance, by a recipient mammal, of a graft from a donor mammal of the same species. The method includes:
administering to the recipient, an inhibitor, e.g., a blocker, of a costimulatory pathway, e.g., a blocker of the CD28-B7 interaction;
introducing, e.g., by intravenous injection, into the recipient mammal, hematopoietic stem cells, e.g., a bone marrow preparation; and
preferably, implanting the graft in the recipient. The hematopoietic cells are believed to prepare the recipient for the graft that follows, by inducing tolerance at both the B-cell and T-cell levels.
In preferred embodiments the CD28-B7 interaction is inhibited by administering a soluble ligand or receptor or antibody for the CD28 or B7, e.g., a soluble CTLA4, e.g., a CTLA4 fusion protein, e.g., a CTLA4 immunoglobulin fusion, e.g, CTLA4/Ig. Preferably, the inhibitor binds B7. In preferred embodiments anti-B7-1 and/or anti-B7-2 antibodies are administered.
In preferred embodiments the donor and recipient are both are humans.
In preferred embodiments the method can be practiced without the administration of hematopoietic space-creating irradiation, e.g., whole body irradiation,.
In certain embodiments the method is practiced without T cell depletion or inactivation, e.g., without the administration of thymic irradiation, or anti-T cell antibodies.
In certain embodiments the method is practiced with T cell depletion or inactivation, e.g., by the administration of thymic irradiation, or anti-T cell antibodies.
In certain embodiments the method is practiced with partial T cell depletion or inactivation, e.g., by the administration of thymic irradiation, or anti-T cell antibodies, in such amount to result in partial depletion of recipient T cells.
In preferred embodiments the method includes administering a sufficiently large number of donor hematopoietic cells to the recipient such that, donor stem cells engraft and give rise to mixed chimerism, without space-creating treatment. Thus, in preferred embodiments inhibitors of both pathways and a quantity of hematopoietic stem cells sufficient to give rise to mixed chimerism, without the need for hematopoietic space-creating irradiation, is administered to the recipient. In preferred embodiments the number of donor hematopoietic cells is at least 200%, is at least equal to, or is at least 75, 50, or 25% as great as, the number of bone marrow cells found in an adult of the recipient species. In preferred embodiments the number of donor hematopoietic stem cells is at least 200%, is at least equal to, or is at least 75, 50,or 25% as great as, the number of bone marrow hematopoietic stem cells found in an adult of the recipient species.
In preferred embodiments, mixed chimerism is induced in the recipient and the state of mixed chimerism is formed in the absence of the induction of hematopoietic space, e.g., in the absence of hematopoietic space created by space creating irradiation, e.g., whole body irradiation.
The number of donor cells administered to the recipient can be increased by either increasing the number of stem cells provided in a particular administration or by providing repeated administrations of donor stem cells.
Repeated stem cell administration can promote engraftment, mixed chimerism, and preferably long-term deletional tolerance in graft recipients. Thus, the invention also includes methods in which multiple hematopoietic stem cell administrations are provided to a recipient. Multiple administration can substantially reduce or eliminate the need for hematopoietic space-creating irradiation. Administrations can be given prior to, at the time of, or after graft implantation. In preferred embodiments multiple administrations of stem cells are provided prior to the implantation of a graft. Two, three, four, five, or more administrations can be provided. The period between administrations of hematopoietic stem cells can be varied. In preferred embodiments a subsequent administration of hematopoietic stem cell is provided: at least two days, one week, one month, or six months after the previous administration of stem cells; when the recipient begins to show signs of host lymphocyte response to donor antigen; when the level of chimerism decreases; when the level of chimerism falls below a predetermined value; when the level of chimerism reaches or falls below a level where staining with a monoclonal antibody specific for a donor PBMC antigen is equal to or falls below staining with an isotype control which does not bind to PBMC""s, e.g. when the donor specific monoclonal stains less than 1-2% of the cells; or generally, as is needed to maintain a level of mixed chimerism sufficient to maintain tolerance to donor antigen.
One or more post graft-implantation-administrations of donor stem cells can also be provided to minimize or eliminate the need for irradiation. Post graft administration of hematopoietic stem cells can be provided: at least two days, one week, one month, or six months after the previous administration of stem cells; at least two days, one week, one month, six months, or at any time in the life span of the recipient after the implantation of the graft; when the recipient begins to show signs of rejection, e.g., as evidenced by a decline in function of the grafted organ, by a change in the host donor specific antibody response, or by a change in the host lymphocyte response to donor antigen; when the level of chimerism decreases; when the level of chimerism falls below a predetermined value; when the level of chimerism reaches or falls below a level where staining with a monoclonal antibody specific for a donor PBMC antigen is equal to or falls below staining with an isotype control which does not bind to PBMC""s, e.g. when the donor specific monoclonal stains less than 1-2% of the cells; or generally, as is needed to maintain tolerance or otherwise prolong the acceptance of a graft.
When multiple stem cell administrations are given one or more of the administrations can include a number of donor hematopoietic cells which is at least 200%, is equal to, or is at least 75, 50, or 25% as great as, the number of bone marrow cells found in an adult of the recipient species; include a number of donor hematopoietic stem cells which is at least 200%, is equal to, or is at least 75, 50, or 25% as great as, the number of bone marrow hematopoietic stem cells found in an adult of the recipient species.
Although methods in which blockers of both pathways are administered may usually eliminate the need for other preparative steps, some embodiments include inactivating T cells, preferably graft reactive T cells of the recipient mammal. This can be accomplished, e.g., by introducing into the recipient mammal an antibody capable of binding to T cells of the recipient mammal. The administration of antibodies, or other treatment to inactivate T cells, can be given prior to introducing the hematopoietic stem cells into the recipient mammal or prior to implanting the graft in the recipient.
Monoclonal preparations can be used in the methods of the invention.
Other preferred embodiments include: the step of introducing into the recipient mammal, donor species-specific stromal tissue, preferably hematopoietic stromal tissue, e.g., fetal liver or thymus. In preferred embodiments: the stromal tissue is introduced simultaneously with, or prior to, the hematopoietic stem cells; the hematopoietic stem cells are introduced simultaneously with, or prior to, the antibody.
Although methods in which a blocker is administered may usually eliminate the need for other preparative steps, some embodiments include the inactivation of thymocytes or T cells, which can be performed prior to hematopoietic stem cell or graft transplantation. In preferred embodiments the method includes diminishing or inhibiting thymocyte or T cell activity, preferably the activity of thymic or lymph node T cells by administering to the recipient a short course of an immunosuppressive agent, e.g., a chemical or drug, e.g., cyclosporine, sufficient to inactivate thymocytes or T cells, preferably thymic or lymph node T cells. The duration of the short course of immunosuppressive agent is: approximately equal to or less than 30, 40, 50, 60, 120, or 365 days; approximately equal to or less than 8-12 days, preferably about 10 days; approximately equal to or less than two, three, four, five, or ten times the 8-12 or 10 day period. The short course can begin: before or at about the time the treatment to induce tolerance is begun, e.g., at about the time stem cells are introduced into the recipient; on the day the treatment to induce tolerance is begun, e.g., on the day stem cells are introduced into the recipient; within 1, 2, 4, 6, 8, 10, or 30 days before or after the treatment to induce tolerance is begun, e.g., within 1, 2, 4, 6, 8, 10, or 30 days before or after stem cells are introduced into the recipient. The short course of an immunosuppressive can be administered in conjunction with an anti-T cell antibody The short course of an immunosuppressive should be sufficient in concentration and duration to inactivate T cells, e.g., thymic or lymph node T cells, which would not be inactivated by antibody-based inactivation of T cells, e.g., inactivation by intravenous administrations of ATG antibody, or similar, preparations.
Although methods in which a blocker is administered may usually eliminate the need for other preparative steps, some embodiments include (optionally): the step of, prior to hematopoietic stem cell transplantation, creating hematopoietic space, e.g., by irradiating the recipient mammal with low dose, e.g., less than 400, preferably less than 300, more preferably less than 200 or 100 cGy, whole body irradiation to deplete or partially deplete the bone marrow of the recipient. As is discussed herein this treatment can be reduced or entirely eliminated. Other methods of creating hematopoietic space, e.g., administering hematopoietic space creating antibodies or drugs, e.g., cyclophosphamide or busulfan, to the recipient, can be used. E.g., hematopoietic space can be formed by administering an inhibitor of cell proliferation, e.g., DSG, or an anti-metabolite, e.g. brequinar, or an anti-T cell antibody, e.g., one or both of an anti-CD4 or anti-CD8 antibody.
In preferred embodiments, the method includes the step of introducing into the recipient a graft obtained from the donor which is obtained from a different organ than the hematopoietic stem cells, e.g., a heart, pancreas, liver, or kidney.
In a particularly preferred embodiment the method includes:
administering to a human recipient an inhibitor, e.g., a blocker, the CD28-B7 interaction;
introducing, e.g., by intravenous injection, into the recipient mammal, hematopoietic stem cells, e.g., a bone marrow preparation; and
implanting the graft in the recipient
The method can be practiced without T cell depletion or inactivation, with T cell depletion or inactivation, or with partial T cell depletion or inactivation. T cell inactivation can be effected by the administration of thymic irradiation, or anti T cell antibodies.
In preferred embodiments donor tissue, e.g., hematopoietic cells, are depleted, e.g., partially or wholly, of donor T cells.
In a preferred embodiment the administration of costimulatory blockade, and preferably of any needed irradiation or T cell depletion, is administered within 48, more preferably, within 24, hours of implantation of the graft.
In a preferred embodiment, the graft is a human cadaveric graft.
In another aspect, the invention features a method of promoting acceptance, by a recipient mammal, e.g., a primate, e.g., a human, of a graft from a donor mammal of the same species. The method includes:
administering to the recipient, an inhibitor of a costimulatory pathway, e.g., one or both of an inhibitor, e.g., a blocker, of the CD40 ligand-CD40 interaction and an inhibitor, e.g., a blocker, of the CD28-B7 interaction;
introducing, e.g., by intravenous injection, into the recipient mammal, hematopoietic stem cells, e.g., a bone marrow preparation, wherein the number of hematopoietic stem cells is sufficient such that mixed hematopoietic chimerism can be induced without whole body irradiation;
preferably, implanting the graft in the recipient. The hematopoietic cells are believed to prepare the recipient for the graft that follows, by inducing tolerance at both the B-cell and T-cell levels.
In preferred embodiments the CD40 ligand-CD40 interaction is inhibited by administering an antibody or soluble ligand or receptor for the CD40 ligand or CD40, e.g., by administering an anti-CD40L antibody, e.g., 5C8 or an antibody with similar efficacy or an antibody which has an epitope which overlaps the epitope of 5C8. Preferably the inhibitor binds the CD40 ligand.
In embodiments wherein the CD28-B7 interaction is inhibited, it can be inhibited by administering a soluble ligand or receptor or antibody for the CD28 or B7, e.g., a soluble CTLA4, e.g., a CTLA4 fusion protein, e.g., a CTLA4 immunoglobulin fusion, e.g., a CTLA4/Ig. Preferably, the inhibitor binds B7. In preferred embodiments anti-B7-1 and/or anti-B7-2 antibodies are administered.
In preferred embodiments CTLA4-Ig and an anti CD40L antibody are administered.
In preferred embodiments the donor and recipient both are humans.
In preferred embodiments, a blocker of the CD40/CD40L interaction, e.g., an anti-CD40L antibody is administered prior to administration of a blocker of the CD28/B7 interaction, e.g., CTLA4/Ig. The CD40/CD40L blocker can be administered on the day donor tissue is introduced and the CD28/B7 blocker administered 2, 3,4 5 or more days later.
In certain embodiments the method is practiced without T cell depletion or inactivation, e.g., without the administration of thymic irradiation, or anti-T cell antibodies.
In certain embodiments the method is practiced with T cell depletion or inactivation, e.g., by the administration of thymic irradiation, or anti-T cell antibodies.
In certain embodiments the method is practiced with partial T cell depletion or inactivation, e.g., by the administration of thymic irradiation, or anti-T cell antibodies, in such amount to result in partial depletion of recipient T cells.
The method includes administering a sufficiently large number of donor hematopoietic cells to the recipient such that, donor stem cells engraft, give rise to mixed chimerism without space-creating treatment. In preferred embodiments the number of donor hematopoietic cells is at least 200%, is at least equal to, or is at least 75, 50, or 25% as great as, the number of bone marrow cells found in an adult of the recipient species. In preferred embodiments the number of donor hematopoietic stem cells is at least 200%, is at least equal to, or is at least 75, 50, or 25% as great as, the number of bone marrow hematopoietic stem cells found in an adult of the recipient species.
In preferred embodiments, mixed chimerism is induced in the recipient and the state of mixed chimerism is formed in the absence of the induction of hematopoietic space, e.g., in the absence of hematopoietic space created by space creating irradiation, e.g., whole body irradiation.
The number of donor cells administered to the recipient can be increased by either increasing the number of stem cells provided in a particular administration or by providing repeated administrations of donor stem cells.
Repeated stem cell administration can promote engraftment, mixed chimerism, and preferably long-term deletional tolerance in graft recipients. Thus, the invention also includes methods in which multiple hematopoietic stem cell administrations are provided to a recipient. Multiple administration can substantially reduce or eliminate the need for hematopoietic space-creating irradiation. Administrations can be given prior to, at the time of, or after graft implantation. In preferred embodiments multiple administrations of stem cells are provided prior to the implantation of a graft. Two, three, four, five, or more administrations can be provided. The period between administrations of hematopoietic stem cells can be varied. In preferred embodiments a subsequent administration of hematopoietic stem cell is provided: at least two days, one week, one month, or six months after the previous administration of stem cells; when the recipient begins to show signs of host lymphocyte response to donor antigen; when the level of chimerism decreases; when the level of chimerism falls below a predetermined value; when the level of chimerism reaches or falls below a level where staining with a monoclonal antibody specific for a donor PBMC antigen is equal to or falls below staining with an isotype control which does not bind to PBMC""s, e.g. when the donor specific monoclonal stains less than 1-2% of the cells; or generally, as is needed to maintain a level of mixed chimerism sufficient to maintain tolerance to donor antigen.
One or more post graft-implantation-administrations of donor stem cells can also be provided to minimize or eliminate the need for irradiation. Post graft administration of hematopoietic stem cells can be provided: at least two days, one week, one month, or six months after the previous administration of stem cells; at least two days, one week, one month, six months, or at any time in the life span of the recipient after the implantation of the graft; when the recipient begins to show signs of rejection, e.g., as evidenced by a decline in function of the grafted organ, by a change in the host donor specific antibody response, or by a change in the host lymphocyte response to donor antigen; when the level of chimerism decreases; when the level of chimerism falls below a predetermined value; when the level of chimerism reaches or falls below a level where staining with a monoclonal antibody specific for a donor PBMC antigen is equal to or falls below staining with an isotype control which does not bind to PBMC""s, e.g. when the donor specific monoclonal stains less than 1-2% of the cells; or generally, as is needed to maintain tolerance or otherwise prolong the acceptance of a graft.
When multiple stem cell administrations are given one or more of the administrations can include a number of donor hematopoietic cells which is at least 200%, is equal to, or is at least 75, 50, or 25% as great as, the number of bone marrow cells found in an adult of the recipient species; include a number of donor hematopoletic stem cells which is at least 200%, is equal to, or is at least 75, 50, or 25% as great as, the number of bone marrow hematopoietic stem cells found in an adult of the recipient species.
Although methods in which blockers of both pathways are administered may usually eliminate the need for other preparative steps, some embodiments include inactivating T cells, preferably graft reactive T cells of the recipient mammal. This can be accomplished, e.g., by introducing into the recipient mammal an antibody capable of binding to T cells of the recipient mammal. The administration of antibodies, or other treatment to inactivate T cells, can be given prior to introducing the hematopoietic stem cells into the recipient mammal or prior to implanting the graft in the recipient.
Monoclonal preparations can be used in the methods of the invention.
Other preferred embodiments include: the step of introducing into the recipient mammal, donor species-specific stromal tissue, preferably hematopoietic stromal tissue, e.g., fetal liver or thymus. In preferred embodiments: the stromal tissue is introduced simultaneously with, or prior to, the hematopoietic stem cells; the hematopoietic stem cells are introduced simultaneously with, or prior to, the antibody.
Although methods in which blockers of both pathways are administered may usually eliminate the need for other preparative steps, some embodiments include the inactivation of thymocytes or T cells, which can be performed prior to hematopoietic stem cell or graft transplantation. In preferred embodiments the method includes diminishing or inhibiting thymocyte or T cell activity, preferably the activity of thymic or lymph node T cells by administering to the recipient a short course of an immunosuppressive agent, e.g., a chemical or drug, e.g., cyclosporine, sufficient to inactivate thymocytes or T cells, preferably thymic or lymph node T cells. The duration of the short course of immunosuppressive agent is: approximately equal to or less than 30, 40, 50, 60, 120, or 365 days; approximately equal to or less than 8-12 days, preferably about 10 days; approximately equal to or less than two, three, four, five, or ten times the 8-12 or 10 day period. The short course can begin: before or at about the time the treatment to induce tolerance is begun, e.g., at about the time stem cells are introduced into the recipient; on the day the treatment to induce tolerance is begun, e.g., on the day stem cells are introduced into the recipient; within 1, 2, 4, 6, 8, 10, or 30 days before or after the treatment to induce tolerance is begun, e.g., within 1, 2, 4, 6, 8, 10, or 30 days before or after stem cells are introduced into the recipient. The short course of an immunosuppressive can be administered in conjunction with an anti-T cell antibody The short course of an immunosuppressive should be sufficient in concentration and duration to inactivate T cells, e.g., thymic or lymph node T cells, which would not be inactivated by antibody-based inactivation of T cells, e.g., inactivation by intravenous administrations of ATG antibody, or similar, preparations.
Although methods in which blockers of both pathways are administered may usually eliminate the need for other preparative steps, some embodiments include (optionally): the step of, prior to hematopoietic stem cell transplantation, creating hematopoietic space, e.g., by irradiating the recipient mammal with low dose, e.g., less than 400, preferably less than 300, more preferably less than 200 or 100 cGy, whole body irradiation to deplete or partially deplete the bone marrow of the recipient. As is discussed herein this treatment can be reduced or entirely eliminated. Other methods of creating hematopoietic space, e.g., administering hematopoietic space creating antibodies or drugs, e.g., cyclophosphamide or busulfan, to the recipient, can be used. E.g., hematopoietic space can be formed by administering an inhibitor of cell proliferation, e.g., DSG, or an anti-metabolite, e.g. brequinar, or an anti-T cell antibody, e.g., one or both of an anti-CD4 or anti-CD8 antibody.
Other preferred embodiments include those in which: the same mammal is the donor of one or both the graft and the hematopoietic cells; and the antibody is an anti-human thymocyte polyclonal anti-serum, obtained, e.g., from a horse or pig.
In preferred embodiments, the method includes the step of introducing into a human recipient, a graft obtained from the donor which is obtained from a different organ than the hematopoietic stem cells, e.g., a heart, pancreas, liver, or kidney.
In a particularly preferred embodiment the method includes:
administering to a human recipient, a blocker of the CD40 ligand-CD40 interaction (optionally, a blocker of the CD28-B7 interaction can also be administered);
introducing, e.g., by intravenous injection, into the recipient mammal, hematopoietic stem cells, e.g., a bone marrow preparation; and
implanting the graft in the recipient.
The method can be practiced without T cell depletion or inactivation, with T cell depletion or inactivation, or with partial T cell depletion or inactivation. T cell inactivation can be effected by the administration of thymic irradiation, or anti T cell antibodies.
In preferred embodiments donor tissue, e.g., hematopoietic cells, are depleted, e.g., partially or wholly, of donor T cells.
In a preferred embodiment the administration of costimulatory blockade, and preferably of any needed irradiation or T cell depletion, is administered within 48, more preferably, within 24, hours of implantation of the graft.
In another aspect, the invention features, a method of promoting acceptance by a recipient mammal, e.g., a primate, e.g., a human, of a graft from a donor mammal. The method includes:
administering to the recipient, an inhibitor, e.g., a blocker, of a costimulatory pathway, (e.g., one or both of, an inhibitor, e.g., a blocker, of the CD40 ligand-CD40 interaction and an inhibitor, e.g., a blocker, of the CD28-B7 interaction);
prior to or simultaneous with transplantation of the graft, introducing into the recipient mammal, donor thymic tissue, e.g., thymic epithelium, preferably fetal or neonatal thymic tissue; and (optionally) implanting the graft in the recipient. The thymic tissue prepares the recipient for the graft that follows, by inducing immunological tolerance at the T-cell level.
In preferred embodiments the CD40 ligand-CD40 interaction is inhibited by administering an antibody or soluble ligand or receptor for the CD40 ligand or CD40, e.g., by administering an anti CD40L antibody, e.g., 5C8 or an antibody with similar efficacy. Preferably the inhibitor binds the CD40 ligand.
In preferred embodiments the CD28-B7 interaction is inhibited by administering a soluble ligand or receptor or antibody for the CD28 or B7, e.g., a soluble CTLA4, e.g., a CTLA4 fusion protein, e.g., a CTLA4 immunoglobulin fusion, e.g, CTLA4/Ig. Preferably, the inhibitor binds B7. In preferred embodiments anti B7-1 and/or anti B7-2 antibodies are administered.
In preferred embodiments CTLA4-Ig and an antiCD40L antibody are administered.
In preferred embodiments one or more of a soluble fragment of CD40 or a soluble fragment of CD28 is administered.
In preferred embodiments the donor and recipient are from the same species, e.g., both are humans.
In preferred embodiments, a blocker of the CD40/CD40L interaction, e.g., an anti-CD40L antibody, is administered prior to administration of a blocker of the CD28/B7 interaction, e.g., CTLA4/Ig. The CD40/CD40L blocker can be administered on the day donor tissue is introduced and the CD28B7 blocker administered 2, 3, 4, 5 or more days later.
In preferred embodiments the recipient is of a first species and the donor is of a second species.
The recipient mammal can be, by way of example, a human. The donor mammal can be, by way of example, a swine, e.g., a miniature swine. The graft is preferably from a discordant species. The graft preferably expresses a major histocompatibility complex (MHC) antigen, preferably a class II antigen. In particularly preferred embodiments the recipient is a primate, e.g., a human, and the donor is a swine, e.g., a miniature swine. antibodies.
In preferred embodiments the method can be practiced without the administration of hematopoietic space-creating irradiation, e.g., whole body irradiation.
In certain embodiments the method is practiced without T cell depletion or inactivation, e.g., without the administration of thymic irradiation, or anti-T cell antibodies.
In certain embodiments the method is practiced with T cell depletion or inactivation, e.g., by the administration of thymic irradiation, or anti-T cell antibodies.
In certain embodiments the method is practiced with partial T cell depletion or inactivation, e.g., by the administration of thymic irradiation, or anti-T cell antibodies, in such amount to result in partial depletion of recipient T cells.
Although methods in which blockers of both pathways are administered may usually minimize or eliminate the need for other preparative steps, some embodiments include inactivating natural killer cells, preferably graft reactive or xenoreactive, e.g., swine reactive, NK cells, of the recipient mammal. This can be accomplished, e.g., by introducing into the recipient mammal an antibody capable of binding to natural killer cells of the recipient mammal, e.g., an anti-CD2 antibody, e.g., MEDI-507. The administration of antibodies, or other treatment to inactivate natural killer cells, can be given prior to introducing the thymic tissue into the recipient mammal or prior to implanting the graft in the recipient. This antibody can be the same or different from an antibody used to inactivate T cells.
Although methods in which blockers of both pathways are administered may usually minimize or eliminate the need for other preparative steps, some embodiments include inactivating T cells, preferably graft reactive or xenoreactive, e.g., swine reactive, T cells of the recipient mammal. This can be accomplished, e.g., by introducing into the recipient mammal an antibody capable of binding to T cells of the recipient mammal. The administration of antibodies, or other treatment to inactivate T cells, can be given prior to introducing the thymic tissue into the recipient mammal or prior to implanting the graft in the recipient. This antibody can be the same or different from an antibody used to inactivate natural killer cells.
One source of anti-NK antibody is anti-human thymocyte polyclonal anti-serum. Preferably, a second anti-mature T cell antibody can be administered as well, which lyses T cells as well as NK cells. Lysing T cells is advantageous for both thymic tissue and graft survival. Anti-T cell antibodies are present, along with anti-NK antibodies, in anti-thymocyte anti-serum. Repeated doses of antibodies, e.g., anti-NK or anti-T cell antibodies, may be preferable. Monoclonal preparations can be used in the methods of the invention.
Although methods in which blockers of both pathways are administered may usually minimize or eliminate the need for other preparative steps, some embodiments include (optionally): the step of, prior to hematopoietic stem cell transplantation, creating hematopoietic space, e.g., by irradiating the recipient mammal with low dose, e.g., less than 400, preferably less than 300, more preferably less than 200 or 100 cGy, whole body irradiation to deplete or partially deplete the bone marrow of the recipient. As is discussed herein this treatment can be reduced or entirely eliminated. Other methods of creating hematopoietic space, e.g., administering hematopoietic space creating antibodies or drugs, e.g., cyclophosphamide or busulfan, to the recipient, can be used. E.g., hematopoietic space can be formed by administering an inhibitor of cell proliferation, e.g., DSG, or an anti-metabolite, e.g. brequinar, or an anti-T cell antibody, e.g., one or both of an anti-CD4 or anti-CD8 antibody.
Other preferred embodiments include: the step of, preferably prior to hematopoietic or thymic tissue transplantation, depleting natural antibodies from the blood of the recipient mammal. Depletion can be achieved, by way of example, by contacting the recipients blood with an epitope which absorbs preformed anti-donor antibody. The epitope can be coupled to an insoluble substrate and provided, e.g., as an affinity column. E.g., an xcex11-3 galactose linkage epitope-affinity matrix, e.g., matrix bound linear B type VI carbohydrate, can be used to deplete natural antibodies. Depletion can also be achieved by hemoperfusing an organ, e.g., a liver or a kidney, obtained from a mammal of the donor species. (In organ hemoperfusion antibodies in the blood bind to antigens on the cell surfaces of the organ and are thus removed from the blood.)
Other preferred embodiments include those in which: the same mammal of the second species is the donor of one or both the graft and the hematopoietic cells; and the antibody is an anti-human thymocyte polyclonal anti-serum, obtained, e.g., from a horse or pig.
In preferred embodiments, the method includes the step of introducing into the recipient a graft obtained from the donor which is obtained from a different organ than the hematopoietic stem cells, e.g., a heart, pancreas, liver, or kidney.
In preferred embodiments the host or recipient is a post-natal individual, e.g., an adult, or a child.
In preferred embodiments the method further includes the step of identifying a host or recipient which is in need of a graft.
In preferred embodiments donor tissue, e.g., thymic tissue, is depleted, e.g., wholly or partially depleted of donor T cells.
In a preferred embodiment the administration of costimulatory blockade, and preferably of any needed irradiation or T cell depletion, is administered within 48, more preferably, within 24, hours of implantation of the graft.
xe2x80x9cPartial T cell depletionxe2x80x9d, as used herein, refers to a condition in which some, but not all, of the subject""s T cells are deleted. In some regimens the administration of a single dose of a T cell depleting agent is useful for creating partial T cell depletion.
A xe2x80x9ccostimulatory pathwayxe2x80x9d, as used herein, is a pathway having a molecule on the surface of a T cell and having a molecule on the surface of an antigen presenting cell. An interaction between these two molecules, in the context of antigen specific recognition by the T cell of an antigen presented by the antigen presenting cell, promotes T cell activation. Examples of costimulatory pathways include the CD40-CD40 ligand pathway and the B7-CD28 pathway.
xe2x80x9cDiscordant species combinationxe2x80x9d, as used herein, refers to two species in which hyperacute rejection occurs when a graft is grafted from one to the other. Generally, discordant species are from different orders, while non-discordant species are from the same order. For example, rats and mice are non-discordant concordant species. Concordant species combinations do not exhibit hyperacute rejection.
xe2x80x9cGraftxe2x80x9d, as used herein, refers to a body part, organ, tissue, or cells. Organs such as liver, kidney, heart or lung, or other body parts, such as bone or skeletal matrix, tissue, such as skin, intestines, endocrine glands, or progenitor stem cells of various types, are all examples of grafts.
xe2x80x9cHematopoietic stem cellxe2x80x9d, as used herein, refers to a cell, e.g., a bone marrow cell, or a fetal liver or spleen cell, which is capable of developing into all myeloid and lymphoid lineages and by virtue of being able to self-renew can provide long term hematopoietic reconstitution. Purified preparations of hematopoietic cells or preparations, such as bone marrow, which include other cell types, can be used in methods of the invention. Although not wishing to be bound by theory, it is believed that the hematopoietic stem cells home to a site in the recipient mammal. The preparation should include immature cells, i.e., undifferentiated hematopoietic stem cells; these desired cells can be separated out of a preparation or a complex preparation can be administered. E.g., in the case of bone marrow stem cells, the desired primitive cells can be separated out of a preparation or a complex bone marrow sample including such cells can be used. Hematopoietic stem cells can be from fetal, neonatal, immature or mature animals. Stem cells derived from the cord blood of the recipient or the donor can be used in methods of the invention. See U.S. Pat. No. 5,192,553, hereby incorporated by reference, and U.S. Pat. No. 5,004,681, hereby incorporated by reference. Donor peripheral hematopoietic stem cells are preferred in some embodiments.
xe2x80x9cImmunosuppressive agent capable of inactivating thymic or lymph node T cellsxe2x80x9d, as used herein, is an agent, e.g., a chemical agent, e.g., a drug, which, when administered at an appropriate dosage, results in the inactivation of thymic or lymph node T cells. Examples of such agents are cyclosporine, FK-506, and rapamycin. Anti-T cell antibodies can also be used. An agent should be administered in sufficient dose to result in significant inactivation of thymic or lymph node T cells which are not inactivated by administration of an anti-T cell antibody, e.g., an anti-ATG preparation. Putative agents, and useful concentrations thereof, can be prescreened by in vitro or in vivo tests, e.g., by administering the putative agent to a test animal, removing a sample of thymus or lymph node tissue, and testing for the presence of active T cells in an in vitro or in vivo assay. Such prescreened putative agents can then be further tested in transplant assays.
xe2x80x9cThymic or lymph node or thymocytes or T cellxe2x80x9d, as used herein, refers to thymocytes or T cells which are resistant to inactivation by traditional methods of T cell inactivation, e.g., inactivation by a single intravenous administration of anti-T cell antibodies, e.g., antibodies, e.g., ATG preparation. xe2x80x9cThymic irradiationxe2x80x9d, as used herein, refers to a treatment in which at least half, and preferably at least 75, 90, or 95% of the administered irradiation is targeted to the thymus. Whole body irradiation, even if the thymus is irradiated in the process of delivering the whole body irradiation, is not considered thymic irradiation.
xe2x80x9cMHC antigenxe2x80x9d, as used herein, refers to a protein product of one or more MHC genes; the term includes fragments or analogs of products of MHC genes which can evoke an immune response in a recipient organism. Examples of MHC antigens include the products (and fragments or analogs thereof) of the human MHC genes, i.e., the HLA genes. MHC antigens in swine, e.g., miniature swine, include the products (and fragments and analogs thereof) of the SLA genes, e.g., the DRB gene.
xe2x80x9cMiniature swinexe2x80x9d, as used herein, refers to a wholly or partially inbred pig. Preferable the swine is from a herd with a coefficient of inbreeding of at least 0.80. The miniature swine can be about 150-250, more preferably, about 200 pounds in weight.
xe2x80x9cHematopoietic space-creating irradiationxe2x80x9d, as used herein, refers to irradiation directed to the hematopoietic tissue, i.e., to tissue in which stem cells are found, e.g., the bone marrow. It is of sufficient intensity to kill or inactivate a substantial number of hematopoietic cells. It is often given as whole body irradiation.
xe2x80x9cThymic spacexe2x80x9d as used herein, is a state created by a treatment that facilitates the migration to and/or development in the thymus of donor hematopoietic cells of a type which can delete or inactivate host thymocytes that recognize donor antigens. It is believed that the effect is mediated by elimination of host cells in the thymus.
xe2x80x9cShort course of a immunosuppressive agentxe2x80x9d, as used herein, means a transitory non-chronic course of treatment. The treatment should begin before or at about the time the treatment to induce tolerance is begun, e.g., at about the time, xenogeneic, allogeneic, genetically engineered syngeneic, or genetically engineered autologous stem cells are introduced into the recipient. e.g., the short course can begin on the day the treatment to induce tolerance is begun, e.g., on the day, xenogeneic, allogeneic, genetically engineered syngeneic, or genetically engineered autologous stem cells are introduced into the recipient or the short course can begin within 1, 2, 4, 6, 8, or 10 days before or after the treatment to induce tolerance is begun, e.g., within 1, 2, 4, 6, 8, or 10 days before or after xenogeneic, allogeneic, genetically engineered syngeneic, or genetically engineered autologous stem cells are introduced into the recipient. The short course can last for: a period equal to or less than about 8-12 days, preferably about 10 days, or a time which is approximately equal to or is less than two, three, four, five, or ten times the 8-12 or 10 day period. Optimally, the short course lasts about 30 days. The dosage should be sufficient to maintain a blood level sufficient to inactivate thymic or lymph node T cells. A dosage of approximately 15 mg/kg/day has been found to be effective in primates.
xe2x80x9cStromal tissuexe2x80x9d, as used herein, refers to the supporting tissue or matrix of an organ, as distinguished from its functional elements or parenchyma.
xe2x80x9cPromoting acceptance of a graftxe2x80x9d as used herein, refers to any of increasing the time a graft is accepted or is functional or decreasing the recipients immune response to the graft, e.g., by the induction of tolerance.
xe2x80x9cTolerancexe2x80x9d, as used herein, refers to an inhibition of a graft recipient""s immune response which would otherwise occur, e.g., in response to the introduction of a nonself MHC antigen into the recipient. Tolerance can involve humoral, cellular, or both humoral and cellular responses. Tolerance, as used herein, refers not only to complete immunologic tolerance to an antigen, but to partial immunologic tolerance, i.e., a degree of tolerance to an antigen which is greater than what would be seen if a method of the invention were not employed. Tolerance, as used herein, refers to a donor antigen-specific inhibition of the immune system as opposed to the broad spectrum inhibition of the immune system seen with immunosuppressants.
xe2x80x9cA blockerxe2x80x9d as used herein, refers to a molecule which binds a member of a ligand/counter-ligand pair and inhibits the interaction between the ligand and counter-ligand or which disrupts the ability of the bound member to transduce a signal. The blocker can be an antibody (or fragment thereof) to the ligand or counter ligand, a soluble ligand (soluble fragment of the counter ligand), a soluble counter ligand (soluble fragment of the counter ligand), or other protein, peptide or other molecule which binds specifically to the counter-ligand or ligand, e.g., a protein or peptide selected by virtue of its ability to bind the ligand or counter ligand in an affinity assay, e.g., a phage display system.
The use of the article xe2x80x9caxe2x80x9d or xe2x80x9canxe2x80x9d non limiting with regard to number except where clearly indicated to be limited by the context. E.g., methods which include administering xe2x80x9canxe2x80x9d inhibitor can include administering one or more than one inhibitor.
Methods of the invention minimize or eliminate the need for hematopoietic space-creating treatment, e.g., irradiation, in many methods of tolerance induction.
In methods of the invention, the creation of thymic space, e.g., by thymic irradiation, the inactivation of recipient peripheral T cells and thymocytes, and the administration of a sufficiently large number of xenogeneic or allogeneic donor stem cells allows the induction of tolerance without subjection the recipient to WBI. The induction of thymic space can reduce the level of donor reactive thymocytes but additional steps (described herein) can be added to further diminish donor thymocyte reactivity.
The invention provides a reliable, non-toxic method of inducing transplantation tolerance. It minimizes the problems of chronic organ graft rejection and immunosuppression-related toxicity. BMT with CTLA4Ig plus MR1 specifically minimizes or eliminates donor-reactive T cells, while avoiding the non-specific depletion or suppression of T cells, which is a component of clinically available immunosuppressive strategies, and can lead to severe complications. This treatment protocol is suitable for both cadaveric and living-related organ transplantation, as it allows the reliable induction of deletional tolerance with a non-toxic conditioning regimen beginning on the day of transplantation. Since the peripheral T cell repertoire is not globally depleted by the conditioning and only a low, minimally myelosuppressive dose of whole body irradiation is given, the clinical usefulness of this approach is extraordinarily high.
Other features and advantages of the invention will be apparent from the following detailed description, and from the claims.